PROJECT SUMMARY/ABSTRACT Malaria, caused by Plasmodium parasites, continues to be a major global health problem, with more than 200 million new infections and nearly 500 thousand deaths annually. Infection initiates when sporozoite stages are inoculated into the skin by the mosquito vector. Sporozoites then move in the skin tissue, enter the blood stream and reach the liver. Sporozoites traverse cells before eventually infecting a hepatocyte within a replication-permissive parasitophorous vacuole (PV). Ensconced in the PV membrane (PVM), a single sporozoite will transmogrify into a liver stage that replicates and then forms tens of thousands of merozoites. These are released from the liver and infect and replicate in blood cells, which causes all clinical symptoms of malaria and enables further parasite transmission. Targeting the pre-erythrocytic sporozoites thus represents an attractive intervention point since they are small in number, and successful disruption of the molecular interactions required for infection would completely prevent disease and transmission. However, little is known about the critical molecular interactions leading to productive sporozoite entry into the host, infection of the liver and invasion of hepatocytes. In this proposal, using novel approaches, we will focus on elucidating the molecular map of infection that involves the sporozoite protein complexes containing P52, P36 and the TRAP complex, and their interactions with the host receptors EphA2, CD81 and PDGFR?, respectively. Currently we only know that P36 and P52 are essential for establishing productive hepatocyte infection with a PVM and that the hepatocyte surface receptors EphA2 and CD81 are critical for PVM formation as well. In Aim 1, we will investigate the dynamics and conditions of release of P52 and P36 from sporozoites during interaction with hepatocytes. Following our preliminary isolation of a P52/P36 complex, we will proceed to determine the precise molecular interactions within this complex and its interaction with EphA2, CD81 and any other novel putative receptor(s) identified. In Aim 2, we will focus on the hepatocyte surface to further study the functional significance of EphA2 during hepatocyte invasion and PV formation as well as its relationship to CD81 and novel receptors. In Aim 3, we will investigate a novel parasite-host interaction pair identified by us between host protein PDGFR? and the sporozoite adhesin TRAP, and the importance of this interaction for host infection in the context of a large TRAP-containing sporozoite protein complex that we have recently identified. Upon completion of the proposed work we will have a clear molecular map of the interactions of critical sporozoite invasion-related protein complexes and host receptor complexes. This data will be important to design targeted interventions such as infection-blocking antibodies or small molecule inhibitors and as such will contribute to the goal of preventing malaria infection.